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Plasmid Files

pOG44

Vector for constitutive expression of a thermolabile Flp recombinase (flp-F70L) in mammalian cells, for use with the Flp-In™ system.

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pOG44 Sequence and MappOG44.dna
Map and Sequence File   
Sequence Author:  Ben Glick
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 NaeI (5455) NgoMIV (5453) DraIII (5352) XmnI (4810) NmeAIII (4359) BpmI (4281) AhdI (4211) AlwNI (3738) PciI (3325) BspQI - SapI (3209) ApaI (2925) PspOMI (2921) Bpu10I (182) NruI (210) MluI (230) SnaBI (592) NcoI - StyI (612) XbaI (896) PflMI (926) BglII (1106) AleI (1123) Bsu36I (1236) BstAPI (1301) EcoRV (1546) SwaI (1810) BspDI * - ClaI * (1917) BbsI (2128) BclI * (2254) NsiI (2290) XcmI (2346) BmgBI (2556) BsmI (2652) AfeI (2752) KasI (2803) NarI (2804) SfoI (2805) PluTI (2807) SphI (2821) EcoNI (2881) SalI (2906) pOG44 5785 bp
NaeI  (5455)
1 site
G C C G G C C G G C C G

Efficient cleavage requires at least two copies of the NaeI
recognition sequence.
NgoMIV  (5453)
1 site
G C C G G C C G G C C G

Efficient cleavage requires at least two copies of the NgoMIV
recognition sequence.
DraIII  (5352)
1 site
C A C N N N G T G G T G N N N C A C

Sticky ends from different DraIII sites may not be compatible.
XmnI  (4810)
1 site
G A A N N N N T T C C T T N N N N A A G
NmeAIII  (4359)
1 site
G C C G A G ( N ) 18-19 N N C G G C T C ( N ) 18-19

Efficient cleavage requires at least two copies of the NmeAIII
recognition sequence.
Sticky ends from different NmeAIII sites may not be compatible.
For full activity, add fresh S-adenosylmethionine (SAM).
BpmI  (4281)
1 site
C T G G A G ( N ) 14 N N G A C C T C ( N ) 14

Efficient cleavage requires at least two copies of the BpmI
recognition sequence.
Sticky ends from different BpmI sites may not be compatible.
After cleavage, BpmI can remain bound to DNA and alter its
electrophoretic mobility.
BpmI quickly loses activity at 37°C.
AhdI  (4211)
1 site
G A C N N N N N G T C C T G N N N N N C A G

The 1-base overhangs produced by AhdI may be hard to ligate.
Sticky ends from different AhdI sites may not be compatible.
AlwNI  (3738)
1 site
C A G N N N C T G G T C N N N G A C

Sticky ends from different AlwNI sites may not be compatible.
PciI  (3325)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
BspQI  (3209)
1 site
G C T C T T C N C G A G A A G N N N N

Sticky ends from different BspQI sites may not be compatible.
SapI  (3209)
1 site
G C T C T T C N C G A G A A G N N N N

Sticky ends from different SapI sites may not be compatible.
SapI gradually settles in solution, so a tube of SapI should be
mixed before removing an aliquot.
ApaI  (2925)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
PspOMI  (2921)
1 site
G G G C C C C C C G G G
Bpu10I  (182)
1 site
C C T N A G C G G A N T C G

Efficient cleavage requires at least two copies of the Bpu10I
recognition sequence.
This recognition sequence is asymmetric, so ligating sticky ends
generated by Bpu10I will not always regenerate a Bpu10I site.
Sticky ends from different Bpu10I sites may not be compatible.
NruI  (210)
1 site
T C G C G A A G C G C T
MluI  (230)
1 site
A C G C G T T G C G C A
SnaBI  (592)
1 site
T A C G T A A T G C A T
NcoI  (612)
1 site
C C A T G G G G T A C C
StyI  (612)
1 site
C C W W G G G G W W C C

Sticky ends from different StyI sites may not be compatible.
XbaI  (896)
1 site
T C T A G A A G A T C T
PflMI  (926)
1 site
C C A N N N N N T G G G G T N N N N N A C C

Sticky ends from different PflMI sites may not be compatible.
BglII  (1106)
1 site
A G A T C T T C T A G A
AleI  (1123)
1 site
C A C N N N N G T G G T G N N N N C A C
Bsu36I  (1236)
1 site
C C T N A G G G G A N T C C

Sticky ends from different Bsu36I sites may not be compatible.
BstAPI  (1301)
1 site
G C A N N N N N T G C C G T N N N N N A C G

Sticky ends from different BstAPI sites may not be compatible.
EcoRV  (1546)
1 site
G A T A T C C T A T A G

EcoRV is reportedly more prone than its isoschizomer Eco32I to
delete a base after cleavage.
SwaI  (1810)
1 site
A T T T A A A T T A A A T T T A

SwaI is typically used at 25°C, but is 50% active at 37°C.
BspDI  (1917)
1 site
A T C G A T T A G C T A
* Blocked by Dam methylation.
ClaI  (1917)
1 site
A T C G A T T A G C T A
* Blocked by Dam methylation.
BbsI  (2128)
1 site
G A A G A C N N C T T C T G N N ( N ) 4

Sticky ends from different BbsI sites may not be compatible.
BbsI gradually loses activity when stored at -20°C.
BclI  (2254)
1 site
T G A T C A A C T A G T
* Blocked by Dam methylation.
BclI is typically used at 50-55°C, but is 50% active at 37°C.
NsiI  (2290)
1 site
A T G C A T T A C G T A
XcmI  (2346)
1 site
C C A N N N N N N N N N T G G G G T N N N N N N N N N A C C

The 1-base overhangs produced by XcmI may be hard to ligate.
Sticky ends from different XcmI sites may not be compatible.
BmgBI  (2556)
1 site
C A C G T C G T G C A G

This recognition sequence is asymmetric, so ligating blunt ends
generated by BmgBI will not always regenerate a BmgBI site.
BsmI  (2652)
1 site
G A A T G C N C T T A C G N

Sticky ends from different BsmI sites may not be compatible.
AfeI  (2752)
1 site
A G C G C T T C G C G A
KasI  (2803)
1 site
G G C G C C C C G C G G
NarI  (2804)
1 site
G G C G C C C C G C G G

Efficient cleavage requires at least two copies of the NarI
recognition sequence.
SfoI  (2805)
1 site
G G C G C C C C G C G G
PluTI  (2807)
1 site
G G C G C C C C G C G G

Efficient cleavage requires at least two copies of the PluTI
recognition sequence.
SphI  (2821)
1 site
G C A T G C C G T A C G
EcoNI  (2881)
1 site
C C T N N N N N A G G G G A N N N N N T C C

The 1-base overhangs produced by EcoNI may be hard to ligate.
Sticky ends from different EcoNI sites may not be compatible.
SalI  (2906)
1 site
G T C G A C C A G C T G
FLP
1202 .. 2473  =  1272 bp
423 amino acids  =  48.6 kDa
Product: site-specific recombinase
FLP is a site-specific recombinase from
Saccharomyces cerevisiae. Recombination occurs at
FRT sequences.
FLP
1202 .. 2473  =  1272 bp
423 amino acids  =  48.6 kDa
Product: site-specific recombinase
FLP is a site-specific recombinase from
Saccharomyces cerevisiae. Recombination occurs at
FRT sequences.
AmpR
4138 .. 4998  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   4138 .. 4929  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
4138 .. 4998  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   4930 .. 4998  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
4138 .. 4998  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
ori
3386 .. 3967  =  582 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
3386 .. 3967  =  582 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
f1 ori
5128 .. 5583  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
f1 ori
5128 .. 5583  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
CMV enhancer
237 .. 616  =  380 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
237 .. 616  =  380 bp
human cytomegalovirus immediate early enhancer
chimeric intron
932 .. 1161  =  230 bp
chimera between introns from adenovirus and
immunoglobulin heavy chain genes
chimeric intron
932 .. 1161  =  230 bp
chimera between introns from adenovirus and
immunoglobulin heavy chain genes
CMV promoter
617 .. 820  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
CMV promoter
617 .. 820  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
SV40 poly(A) signal
2610 .. 2731  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
2610 .. 2731  =  122 bp
SV40 polyadenylation signal
AmpR promoter
4999 .. 5103  =  105 bp
AmpR promoter
4999 .. 5103  =  105 bp
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